SMAD2 Inactivation Inhibits CLDN6 Methylation to Suppress Migration and Invasion of Breast Cancer Cells

Abstract

The downregulation of tight junction protein CLDN6 promotes breast cancer cell migration and invasion; however, the exact mechanism underlying CLDN6 downregulation remains unclear. CLDN6 silence is associated with DNA methyltransferase 1 (DNMT1) mediated DNA methylation, and DNMT1 is regulated by the transforming growth factor beta (TGFβ)/SMAD pathway. Therefore, we hypothesized that TGFβ/SMAD pathway, specifically SMAD2, may play a critical role for CLDN6 downregulation through DNA methyltransferase 1 (DNMT1) mediated DNA methylation. To test this hypothesis, we blocked the SMAD2 pathway with SB431542 in two human breast cancer cell lines (MCF-7 and SKBR-3). Our results showed that treatment with SB431542 led to a decrease of DNMT1 expression and the binding activity for CLDN6 promoter. The methylation level of CLDN6 promoter was decreased, and simultaneously CLDN6 protein expression increased. Upregulation of CLDN6 inhibited epithelial to mesenchymal transition (EMT) and reduced the migration and invasion ability of both MCF-7 and SKBR-3 cells. Furthermore, knocked down of CLDN6 abolished SB431542 effects on suppression of EMT associated gene expression and inhibition of migration and invasion. Thus, we demonstrated that the downregulation of CLDN6 is regulated through promoter methylation by DNMT1, which depends on the SMAD2 pathway, and that CLDN6 is a key regulator in the SMAD2/DNMT1/CLDN6 pathway to inhibit EMT, migration and invasion of breast cancer cells.

Knockdown of CLDN6 inhibits cell proliferation and migration via PI3K/AKT/mTOR signaling pathway in endometrial carcinoma cell line HEC-1-B

“…Our results were in line with previous studies, indicating an oncogenic role of CLDN6. 24 – 26 However, the exact mechanisms remain poor. Significantly, a former report has demonstrated that enhanced proliferation and migration of tumor cells may be caused by the altered change in ionic microenvironments in the lateral membranes.…”

“… 6 – 8 CLDN6 may suppress migration and invasion of cancer cells via EMT inhibition. 26 However, the mechanisms that could explain how CLDN6 regulate cell proliferation rate, migration, and invasiveness still remain to be revealed.…”

Claudins: New Players in Human Fertility and Reproductive System Cancers

“…The overexpression of claudin-6 was showed to inhibit migration and invasion of MCF-7 cells [104]. Lu et al demonstrated that SMAD2 causes downregulation of claudin-6 by DNA (cytosine-5)-methyltransferase 1 (DNMT1)-mediated DNA methylation, which results in increased migration and invasion of breast cancer cells [105]. In addition, research has shown that estrogen receptor β (ERβ) plays an anti-cancer role in breast cancer, and Song et al demonstrated that this effect is mediated by claudin-6 [106].…”

MicroRNA‑27a inhibits trophoblast cell migration and invasion by targeting SMAD2: Potential role in preeclampsia

“…Similar to other SMADs, SMAD2 serves a role in the transmission of extracellular signals from TGF-β ligands into the cell nucleus and mediates cell proliferation, apoptosis and differentiation (45). The TGF-β-SMAD2/3 signaling pathway is involved in epithelial-mesenchymal transition in several malignant tumors, including hepatocellular carcinoma (46), breast cancer (47) and colorectal cancer (48). In addition, SMAD2 also contributes to PE initiation (49).…”